INITIATION OF LAGGING-STRAND DNA SYNTHESIS

滞后链 DNA 合成的起始

• 批准号: 3285795
• 负责人: KENNETH J MARIANS
• 金额: $ 26.09万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-07-01 至 1992-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3285795
• 关键词: DNA binding protein; DNA directed DNA polymerase; DNA footprinting; DNA replication; Escherichia coli; adenosinetriphosphatase; circular DNA; exopeptidase; genetic manipulation; methylation; molecular cloning; nucleic acid sequence; point mutation; proteins; transposon /insertion element

A major part of the effort of this laboratory is directed at developing an understanding of how the replication machinery gains access to the DNA template and subsequently remains bound over tremendous distances while actively synthesizing both the leading- and lagging-strands of DNA. To this end, detailed investigations of the activities of enzymes present at the replication fork are presented. Particular attention is given to the protein-protein and protein-DNA interactions that characterize the replication fork. Studies will be continued that are designed to characterize the interaction of the primosome (a mobile priming apparatus that requires seven E. coli-encoded proteins for assembly and which can assemble at a specific site on the DNA (primosome assembly sites), migrate processively 5' to 3' and occasionally synthesize a primer) with primosome assembly sites. The protein responsible for primosome movement will be established and the three genetically undefined primosomal proteins (replication factor Y (protein n') and proteins n and n") will be molecularly cloned and their genes characterized. The structure and movement of the replication fork will be investigated using novel primer-templates that sustain rolling- circle DNA replication in which synthesis of the leading- and lagging-strands of DNA are coupled. The protein-protein interactions required and the role of the DNA polymerase III holoenzyme in coupling the replication machinery at the fork will be established. Rates of replication fork movement will be measured and factors that influence progress of the replication fork will be identified.

这个实验室的主要工作是 了解复制机制如何获得 进入DNA模板，随后保持结合， 同时积极地综合两个领先的- 和落后的DNA链。 为此，详细调查 存在于复制叉上的酶的活性是 提出了 特别注意蛋白质-蛋白质 和蛋白质与DNA的相互作用 叉子 将继续进行旨在表征 所述原体（一种移动的引发装置， 需要7个E用于组装的大肠杆菌编码的蛋白质， 在DNA上的特定位点组装（原核体组装位点）， 将引物5'迁移到3'，偶尔合成引物） primosome组装位点。 蛋白质负责 原始体运动将建立和三个基因 未定义的primosomal蛋白质（复制因子Y（蛋白质n '）） 以及蛋白质n和n”）将被分子克隆， 表征了 复制叉的结构和运动将 研究使用新的引物模板，维持滚动- 环DNA复制，其中合成的主导和 DNA的滞后链被偶联。 蛋白质-蛋白质 所需的相互作用和DNA聚合酶III的作用 全酶在耦合复制机械叉将 建立。 复制叉移动的速率将为 影响复制进度的因素 叉子将被识别。